The Fc region of an immunoglobulin mediates effector functions that have been divided into two categories. In the first are functions that occur independently of antigen binding; these functions confer persistence in circulation and the ability to be transferred across cellular barriers by transcytosis (see Ward and Ghetie, Therapeutic Immunology 2:77-94, 1995, Capon et al. Nature 1989). The circulatory half-life of the IgG subclass of immunoglobulins is regulated by the affinity of the Fc region for the neonatal Fc receptor for FcRn (Ghetie et al. Nature Biotechnol. 15:637-640, 1997; Kim et. al., Eur. J. Immunol. 24:542-548, 1994; Dall'Acqua et al. (J. Immunol. 169:5171-5180, 2002). The second general category of effector functions include those that operate after an immunoglobulin binds an antigen. In the case of IgG, these functions involve the participation of the complement cascade or Fc gamma receptor (FcγR)-bearing cells. Binding of the Fc region to an FcγR causes certain immune effects, for example, endocytosis of immune complexes, engulfment and destruction of immunoglobulin-coated particles or microorganisms (also called antibody-dependent phagocytosis, or ADCP), clearance of immune complexes, lysis of immunoglobulin-coated target cells by killer cells (called antibody-dependent cell-mediated cytotoxicity, or ADCC), release of inflammatory mediators, regulation of immune system cell activation, and regulation of immunoglobulin production.
While it is possible to generate Fc-containing, heterodimeric polypeptides, current methods require either coexpression of the two heavy chain portions of a heterodimeric Fc region or chemical conjugation of the dimeric Fc region to one or more binding sites (e.g., a Fab domain). Coexpression of these constructs leads to production of complex mixtures representing all possible pairings of starting material in addition to aggregates and inactive protein. Consequently, yields of the desired functional polypeptide are relatively low. Recently, single chain Fc molecules have been developed which overcome many of these problems. These molecules comprise a single chain Fc region in which the component Fc moieties are genetically-fused in a single polypeptide chain such that they form a functional, dimeric Fc region. These single chain polypeptides comprise a linker which is not present in naturally occurring Fc constructs and, therefore, may create unwanted immune responses or may prevent other protein:protein interactions. These single chain Fc constructs may also have lower stability potentially due to constraints imposed by the covalent linker.
Accordingly, there is a need for Fc-containing heterodimeric polypeptides which can be produced efficiently and robustly and which minimize and preferably do not comprise extraneous amino acid sequences.